Project Summary/Abstract Islets of Langerhans are micro-organs of the pancreas comprised of several cell types which secrete hormones which maintain glucose homeostasis. The insulin secreting ?-cells are excitable cells which are sensitive to changes in blood glucose levels, and which metabolize glucose to generate ATP which bind to ATP sensitive potassium channels. Subsequent activation of voltage gated calcium channels increases cytosolic calcium activity ([Ca2+]) which triggers insulin release. Connexin36 (Cx36) gap junctions couple ?- cells by allowing the passage of small cationic molecules which promotes [Ca2+] oscillation synchronization and coordinates the dynamics of insulin secretion, which is necessary for its maximal hypoglycemic affect. There is significant heterogeneity in pancreatic ?-cells in function of Cx36, ion channels, and thus responses to glucose. Studying how this heterogeneity affects coupled electrical behavior is difficult without a means of perturbing individual cells or groups of cells in the islet and studying the response. In order to directly control [Ca2+] dynamics, insulin secretion, and study coupled electrophysiology properties of the islet we have introduced Channelrhodopsin-2 (ChR2, a light activated non-specific cationic channel) into the ?- cell which can generate optically stimulated action potentials. We have created a mouse line with ?-cell specific ChR2 expression and verified that we can optically stimulate spatio-temporally defined [Ca2+] and insulin secretion. To understand the effects of heterogeneity in coupled ?-cell electrophysiology, will test the hypothesis that subpopulations of ?-cells exert disproportionate control over the islet?s electrical dynamics and insulin release and their control is disrupted under diabetic conditions. We will test 2 specific aims; 1) Test for functional subpopulations and quantify how they exert control over each; 2) Examine the role of cytokine induced electrical dysfunction and its effect on subpopulations and coordinated insulin secretion. To study how subpopulations of ?-cell exert control over each other we will define local activation areas using confocal fluorescent microscopy and observe [Ca2+] activity in unstimulated regions. This information will provide spatial maps of excitability throughout the islet and under varying glucose and Cx36 levels. We will test how cytokines alter the ability of subpopulations to regulate control over other regions and how this affects general oscillatory behavior. Elucidating the role of subpopulations in the pancreatic islet and how inflammatory cytokines affect the ability of the subpopulations to control islet behavior could lead to improved knowledge of therapies for diabetes. And will improve general knowledge of coupled ?-cell electrophysiology.